奶牛胰腺对功能性氨基酸的特异性感知与响应网络

The low starch digestibility (35%-60%) in the small intestine of dairy cows is dominated by insufficient pancreatic α-amylase secretion, and it is of great significance to improve the intestinal starch digestion, which is unsolved yet. We have proved the modulation of α-amylase secretion by functional amino acids (FAA), while the potential signaling pathways stay unclear. The existence of sense-transport system in the intestine and pancreas, as well as the highest response of pancreas to FAA, has been recently evidenced in mice. However, few studies were conducted in dairy cows. Hence, we hypothesize that the pancreas presents higher response degree to FAA than other organs, and FAA trigger pancreatic exocrine secretion via a sense-transport-response network in dairy cows. With the applications of isotope-tracer technology and primary cell culture, the objectives of this project are to: 1) explore the regulatory dynamics of FAA (Leu and Phe) on protein synthesis rates among different organs of dairy cows, thus confirm the strongest response degree of pancreas; 2) systematically establish the specific sense-transport-response network of FAA-mediated pancreatic exocrine function. The implementation of this project will provide theoretical basis for energy efficient utilization of dairy cows.

胰腺α-淀粉酶分泌不足是限制奶牛小肠淀粉消化率（35%-60%）的关键因素，提高奶牛小肠淀粉消化率的意义重大，但此问题仍悬而未解。本团队前期已证明，功能性氨基酸可调控胰腺α-淀粉酶分泌，但其感知传导机制并不清楚。初步研究表明，小鼠肠道和胰腺等组织存在功能性氨基酸感知-转运网络，且胰腺响应度最高，而奶牛上鲜有报道。因此，申请者提出假设：“奶牛胰腺对功能性氨基酸的响应高于其它器官，且存在感知-转运-应答网络，通过该网络实现对胰腺外分泌功能的特异性调控”。项目拟通过奶牛活体同位素示踪和原代细胞培养等技术，比较研究功能性氨基酸（亮氨酸、苯丙氨酸）对奶牛不同组织蛋白质合成速率的调控规律，并确证胰腺的高响应度，系统构建胰腺对功能性氨基酸的特异性感知-转运-响应分子网络，为促进奶牛小肠淀粉的高效利用提供理论依据。

通过荷斯坦青年牛、奶山羊的饲养代谢试验，以及胰腺腺泡原代细胞培养试验，以小肠淀粉消化利用、血液指标等为表型指标，系统比较研究了反刍动物胰腺对功能性氨基酸的特异性感知与响应网络。取得了如下结果：. （1）根据淀粉在不同消化道部位消化利用及体内代谢特点，成功构建了通过非手术动物间接评估淀粉在小肠消化利用的预测模型。（2）过瘤胃亮氨酸（RP-Leu）提高荷斯坦青年牛血液葡萄糖水平，不影响全肠道淀粉表观消化率和瘤胃发酵参数，降低粪便中淀粉降解菌的相对丰度，提高粪便中纤维降解菌的丰度，增加粪便pH值，降低丙酸的百分比。（3）与过瘤胃赖氨酸（RP-Lys）相比，添加RP-Leu提高荷斯坦青年牛血液葡萄糖水平、粪便pH，降低丙酸的百分比，提高粪便中纤维降解菌的丰度。（4）RP-Leu和过瘤胃淀粉（RES）水平对青年牛粪便丁酸摩尔百分比和纤维降解菌等关键指标存在交互作用，RES水平越高，RP-Leu提高小肠淀粉消化利用的效果越强。（5）日粮添加RP-Leu提高青年奶山羊和泌乳奶山羊日增重，对泌乳奶山羊产奶量及乳成分无影响。（6）亮氨酸可提高蛋白质合成mTOR信号通路下游关键蛋白因子S6K1和4EBP1磷酸化水平，调节胰腺α-淀粉酶活性；在本研究设置的赖氨酸浓度范围内，赖氨酸对mTOR通路和胰腺α-淀粉酶均未产生影响。. 上述结果证实了项目申请书提出的理论假设“奶牛胰腺存在感知-转运-应答网络，通过该网络实现对胰腺外分泌功能的特异性调控”，即：亮氨酸主要以信号分子形式特异性刺激胰腺mTOR蛋白质合成信号通路，增强下游关键蛋白因子p70S6K和4EBP1的磷酸化水平，促进青年牛和奶山羊胰腺α-淀粉酶合成分泌，提高小肠淀粉消化利用，改善后肠道健康；过瘤胃淀粉流量越高，亮氨酸提高小肠淀粉消化利用的作用效果越显著。本研究为应用功能性氨基酸提高反刍动物消化吸收能力提供了理论依据。